Novel Uses of Willowherb Extract

Abstract

This invention relates to a *Salix viminalis* extract, preferably *Salix frieshes* extract, which can be used as a prebiotic.

Description

[0001] This invention relates to Epilobium extracts, preferably Epilobium Fleischeri extract, for use as prebiotics.

[0002] The skin is the outermost protective layer of an organism and the largest organ in the body. It acts as a barrier, protecting the body from external factors. Furthermore, it is well known that a wide variety of microorganisms reside on the skin surface, forming the skin microbiota (often also called the dermal flora). Resident microbiota are present in the upper part of the epidermis and aggregate within and around hair follicles. Most microorganisms on human skin are bacteria that provide benefits to the host (often called symbionts or good bacteria). These symbionts typically live peacefully with the host, benefiting from their protected ecological niche. However, the microbiome can also include or be exposed to harmful or potentially harmful bacteria (bad bacteria, also known as pathogens), such as *Staphylococcus aureus*, *Corynebacterium tuberculostearicum*, and *Corynebacterium kroppenstedtii*, the latter two of which, as part of the conditionally pathogenic group defined as "corynebacteria," are increasingly associated with major infections. Pathogenic microorganisms can occasionally be harmful to their hosts when present on the skin, especially if their presence exceeds a certain threshold. However, once they enter the body, for example, through ingestion or through skin incisions, these bacteria can clearly become harmful. Therefore, any disruption to the balance between good and bad bacteria presents an opportunity for disease to develop. Thus, achieving and maintaining a balanced microbiome—that is, keeping human surfaces, such as the skin, in a healthy and infection-free state—is often desirable.

[0003] Microbial balance can be achieved, for example, by reducing harmful bacteria through the application of a topical composition containing one or more antimicrobial compounds. However, most, if not all, antimicrobial compounds contained in such topical compositions do not work selectively. In other words, the antimicrobial compounds do not act selectively on only harmful bacteria, but also on beneficial bacteria. Therefore, the use of such topical compositions often leads to the depletion of beneficial bacteria, which is detrimental. Furthermore, such antibiotic compounds may promote highly undesirable antibiotic resistance.

[0004] Bacteria known to have beneficial effects on skin health include Staphylococcus hominis, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium accolens.

[0005] Staphylococcus aureus has been shown to produce hominicin, a bacteriocin that is effective against clinically relevant Staphylococcus aureus strains, including MRSA.

[0006] Staphylococcus epidermidis is a well-known dermal symbiont. Current research has repeatedly confirmed that this bacterium is an important component of a healthy microbiome and also plays a beneficial role in acne by limiting the over-colonization and inflammation of pathogenic Propionibacterium acnes (C. acnes).

[0007] Crowded Corynebacterium can hydrolyze triglycerides on the skin surface in vivo, thereby releasing free fatty acids that fight pneumococcus. Therefore, crowded Corynebacterium plays a beneficial role in shaping the human microbiome.

[0008] *Propionibacterium granulosum* is present as part of the skin microbiota in sebum-rich areas, but its abundance is far lower than that of *Propionibacterium acnes*. Since *Propionibacterium granulosum* co-localizes with *Propionibacterium acnes* in the stratum corneum and hair follicles, it is believed that *Propionibacterium acnes* competes with *Propionibacterium granulosum*. *Propionibacterium granulosum* has been reported to be depleted in the skin microbiota of patients with papulopustular rosacea, suggesting that this species plays an important role in maintaining a normal skin barrier by preventing the growth of potential pathogens. It has also been shown that *Propionibacterium granulosum* can stimulate the host immune system.

[0009] Yunnan Micrococcus is part of the Micrococcus luteus group. Micrococcus luteus is a non-pathogenic species. They can also be used for detoxification or biodegradation of environmental pollutants.

[0010] On the other hand, microorganisms known to have negative effects on skin health include Staphylococcus capitis; Corynebacterium tuberculostearicum; Lawsonella clevelandensis; Staphylococcus aureus; and C. kroppenstedtii.

[0011] Staphylococcus aureus is a well-known skin pathogen primarily associated with the development of atopic dermatitis.

[0012] Staphylococcus cephalosporin is a symbiotic organism that is particularly found on the head and scalp and has an affinity for sebaceous glands. It is also a known opportunistic pathogen and is presumed to be associated with acne formation.

[0013] Corynebacterium tuberculosis is a lipophilic species. Lipophilic Corynebacteria are a particularly relevant subgroup because they are often associated with infections in hospitalized patients and tend to exhibit multidrug resistance to antibiotics. Corynebacterium tuberculosis is believed to play a key role in inflammatory skin conditions because it has been shown to trigger an inflammatory cascade via NF-κB in skin cells.

[0014] Lawnonia clevelandii is a recently discovered bacterium and part of the skin's commensal microbiota. It is commonly found in the nostrils of adults (nasal microbiota) and is likely a low-virulence bacterium. However, routine microbiological approaches may underestimate the incidence of Lawnonia clevelandii infections, as these infections have been observed in the formation of various types of abscesses.

[0015] Corynebacterium cropensteine ​​is considered an opportunistic pathogen and is associated with skin redness, inflammatory skin diseases, and rosacea.

[0016] Based on the different roles and properties of individual microorganisms as part of the microbiome, this creates a need for a well-balanced and healthy composition. Certain mechanisms in the human body contribute to a balanced microbiome. For example, keratinocytes, one of the main cell types abundant on the human body's surface (such as the skin), are known to secrete antimicrobial peptides (a natural defense system), preparing the skin to resist attacks from harmful bacteria.

[0017] However, a balanced gut microbiota can also be achieved or supported by utilizing prebiotics, which promote the growth of beneficial bacteria (relative to harmful bacteria). Therefore, the use of prebiotics leads to an increase in the benefits derived from beneficial bacteria. Beneficial bacteria are said to provide benefits to their hosts, for example, by competing with harmful bacteria for nutrient availability and through the secretion of metabolites.

[0018] Extracts of the genus *Salix*, such as *Salix flecherii* extract, are cosmetic ingredients used to regulate sebum and treat oily skin, and to reduce *Propionibacterium acnes* in vitro. However, little is known about their effects on other microorganisms, particularly skin symbiotic microorganisms and potentially pathogenic microorganisms.

[0019] It has now been surprisingly discovered that the use of *Salix frescoeflorae* extract, when applied to the human body surface, provides a balance to the gut microbiota. It has also been found that this balance is achieved through *Salix frescoeflorae* extract, which acts as a prebiotic, preferentially promoting the growth of good bacteria over bad bacteria.

[0020] The benefits of a balanced microbiome can be achieved through various mechanisms, and not every substance that provides a balanced microbiome necessarily acts as a prebiotic. The invention of this application is achieved by discovering that extracts of the genus *Gynostemma*, such as preferably *Gynostemma flecherum* extract, can function as a prebiotic, which was previously unknown to the inventors.

[0021] Therefore, in a first aspect, the present invention relates to the use of *Salix viminalis* extracts, preferably *Salix flecherii* extracts, as prebiotics when applied to the external surface of the human body.

[0022] The prebiotic effect is characterized by the fact that the *Salix viminalis* extract, preferably *Salix flecheri* extract, promotes the growth of good bacteria in the skin microbiome, such as preferably *Staphylococcus aureus*, *Staphylococcus epidermidis*, *Micrococcus yunnanensis*, *Propionibacterium granulosum*, and *Corynebacterium clavatum* (preferably all of these good bacteria), more than bad bacteria, such as preferably *Staphylococcus capsulatum*, *Corynebacterium tuberculosis*, *L. laurentii*, *S. aureus*, and at least one of *Corynebacterium cropenaeus* (preferably all of these bad bacteria), compared to an untreated control (i.e., a control not treated with *Salix viminalis* extract, preferably *S. flecheri* extract).

[0023] In another aspect, the present invention relates to extracts of the genus *Salix*, preferably *Salix friesiensis* extract, said extract being used to provide a balance of the microbial community when applied to the surface of the human body.

[0024] The resulting microbial balance is achieved through the use of *Salix viminalis* extracts, preferably *Salix friesiensis* extract, as prebiotics, since these extracts have been found to preferentially promote the growth of beneficial bacteria in the skin microbiome, such as at least one of *Staphylococcus aureus*, *Staphylococcus epidermidis*, *Micrococcus yunnanensis*, *Propionibacterium granulosum*, and *Corynebacterium clausti* (preferably all of these beneficial bacteria), exceeding the growth of harmful bacteria, such as at least one of *Staphylococcus capitulata*, *Corynebacterium tuberculosis*, *Lawsonia clevelandii*, *Staphylococcus aureus*, and *Corynebacterium cropensteinii* (preferably all harmful bacteria).

[0025] As used herein, the term "Salix genus extract" encompasses extracts from the flowers, leaves, and / or stems of any Salix genus species, such as preferably Salix friesiensis, Epilobium parviflorum, and / or Epliobium angustifolium.

[0026] Suitable extracts particularly include extracts of *Willow Leaf Flesh* and *Willow Leaf Blue*. Most preferably, in all embodiments of the invention, is *Willow Leaf Flesh* extract having all the definitions and preferences given herein.

[0027] The *Salix fleurone* extract according to the present invention is obtained by extracting the flowers / leaves and stems of the plant and can be used as... -SEBUM was purchased from DSM Nutritional Products.

[0028] Willowherb extract suitable for the purposes of this invention can be used as Defenscalp TM Purchased from Lucas MeyerCosmetics.

[0029] The extract of *Salix viminalis* suitable for use according to the purposes of this invention is commercially available from Kingherbs.

[0030] Preferably, in all embodiments of the invention, the *Salix* extract is prepared by extraction with a polar solvent (e.g., water, ethanol, glycerol, ethylene glycol, or mixtures thereof), preferably with water.

[0031] Preferably, the *Salix* extract according to the invention is an aqueous extract. This *Salix* aqueous extract can be prepared by a method comprising the following steps: grinding aerial parts, such as flowers, leaves, and / or stems of the corresponding *Salix* species, said flowers, leaves, and / or stems preferably pre-dried, more preferably dried in hot air; then extracting the dried plant with a solvent, preferably a mixture of ethanol and water; and then removing the ethanol, for example, by (vacuum) distillation, to obtain the corresponding *Salix* aqueous extract.

[0032] Preferably, in all embodiments of the invention, the *Willowia* extract is an aqueous extract. This *Willowia frescoe* aqueous extract can be prepared by a method comprising the following steps: grinding the aerial parts of *Willowia frescoe*, which are preferably pre-dried, more preferably dried in hot air; extracting the dried plant with a solvent, preferably an ethanol / water mixture; and then removing the ethanol, for example, by (vacuum) distillation, to obtain the *Willowia frescoe* aqueous extract.

[0033] In all embodiments of the invention, the aqueous extract comprises a *Salix viminalis* extract, preferably *Salix frieshes* extract, and may be blended with other ingredients, such as preferably glycerol, citric acid, and preservatives (e.g., potassium sorbate), for example, before being incorporated into the composition according to the invention.

[0034] Willowherb extracts, preferably the Willowherb frescoe extract according to the present invention, provide a balance of the microbial community when applied to the external surface of the human body.

[0035] Exposure to *Gynostemma pentaphyllum* extracts, preferably *Gynostemma flecherum* extract, increases the number of beneficial bacteria, and consequently, the production of metabolites by these beneficial bacteria may also increase. Without being bound by theory, these metabolites are believed to contribute to the body's natural defense system.

[0036] Therefore, in another aspect, the present invention relates to extracts of the genus *Salix*, preferably *Salix flecheri* extract, for the use of improving the natural defense system of the human body's outer surface through microbial community balance.

[0037] Therefore, in another aspect, the present invention relates to a method for improving the natural defense system of the human body surface through the balance of the microbial community, the method comprising the step of applying a willowherb extract, preferably a friesherb extract, to the outer surface of the human body.

[0038] The prebiotic effect according to the present invention is characterized by the fact that at least one microorganism selected from the group consisting of Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium pumilum in the skin microbiota of the individual in need, preferably with increased differential abundance of all these microorganisms.

[0039] The prebiotic effect according to the invention is preferably further characterized by the fact that at least one microorganism selected from the group consisting of Staphylococcus capillaris, Corynebacterium tuberculosis, Lawsonia clevelandii, Staphylococcus aureus, and Corynebacterium cropensteine ​​in the skin microbiome of the individual in need, preferably with reduced differential abundance of all microorganisms.

[0040] Therefore, in another aspect, the present invention also relates to the use of *Salvia splendens* extract, preferably *Salvia splendens freschaeferi* extract, or topical compositions comprising *Salvia splendens* extract, preferably *Salvia splendens freschaeferi* extract, for increasing the differential abundance of at least one microorganism selected from the group consisting of *Staphylococcus aureus*, *Staphylococcus epidermidis*, *Micrococcus yunnanensis*, *Propionibacterium granulosum*, and *Corynebacterium clavatum* in the skin microbiota of an individual in need, preferably all of the microorganisms. Preferably, the use covers at least one microorganism selected from the group consisting of *Staphylococcus capitulata*, *Corynebacterium tuberculosis*, *Lawsonia clevelandii*, *Staphylococcus aureus*, and *Corynebacterium cropensteinii*, preferably with a concomitant decrease in the differential abundance of all microorganisms.

[0041] In another embodiment, the present invention also provides a method for providing protection to the outer surface of the human body by increasing the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium claustrophosum, preferably all of the microorganisms, the method comprising the step of locally applying a Willowleaf extract, preferably a Willowleaf freschaefer extract, to the outer surface.

[0042] As used herein, the term "external surface of the human body" encompasses skin, as well as scalp (including hair and armpits) and oral cavity. Preferably, in all embodiments of the invention, the external surface of the human body treated according to the invention is the skin of the face, neck, and / or body, most preferably the face (including (lateral) cheeks, forehead, nose, and chin).

[0043] As used herein, the phrase "microbial community balance" preferably refers to maintaining the human body surface (e.g., skin, scalp (including hair), armpits, and mouth) in a healthy and infection-free state. Microbial community balance is achieved through the action of *Gynostemma pentaphyllum* extracts, preferably *Gynostemma flecherum* extract, as prebiotics (i.e., preferentially promoting the growth of good bacteria on the human body surface relative to bad bacteria).

[0044] As used in this article, the term "skin microbiota" refers to the microbiota that colonizes a specific area of ​​an individual's skin (e.g., but not limited to the forehead, forearms, cheeks, or scalp).

[0045] As used in this paper, “differential abundance” refers to the logarithmic change in the fold of taxa abundance between two cases. This analytical approach outputs differences, allowing the identification of microbial taxa associated with certain biological or clinical conditions. Differences can be ranked and sorted from lowest to highest. These “rankings” provide information about the relative association of a characteristic with a given covariate (i.e., treatment or time).

[0046] The “differential abundance” used in this paper was determined using software called Songbird, as listed in Morton et al. (Nat. Commun., 2019.10(1): 2719), which is included here by reference. This software allows for the construction of statistical models to test for differences between different treatments and to calculate differences from relative abundance data from 16S rRNA sequencing. The output is a file containing rankings of features for certain metadata categories. The higher the ranking, the greater its relevance to that category (i.e., treatment).

[0047] As used herein, the term “increase or decrease in differential abundance” is defined as an increase or decrease in the differential abundance of a particular microorganism when compared to the differential abundance of a corresponding symmetrical skin region treated with placebo in the same individual.

[0048] In all embodiments of the invention, the *Salix viminalis* extract, preferably *Salix flecherii* extract, is preferably applied in the form of a topical composition comprising an effective amount of the *Salix viminalis* extract, preferably *Salix flecherii* extract, and a cosmetically acceptable carrier.

[0049] The term "effective dose" refers to the amount necessary to achieve the desired physiological effect. The physiological effect can be achieved by a single dose application or by repeated application. Of course, the applied dose can vary depending on known factors (e.g., the physiological characteristics of a particular composition containing *Salix viminalis* extract, preferably *Salix frieshe* extract, and the manner and route of its administration; the recipient's age, health, and weight; the type of concurrent treatment; the frequency of treatment; and the desired effect) and can be adjusted by those skilled in the art.

[0050] Typically, the amount of *Gnaphalium affine* extract in the topical composition according to the invention, preferably *Gnaphalium affine* freschaeferum extract, is selected from the range of 0.001 wt% to 10 wt%, more preferably 0.01 wt% to 8 wt%, even more preferably 0.1 wt% to 7 wt%, further preferably 0.1 wt% to 5 wt%, and even more preferably 0.1 wt% to 3 wt% based on the total weight of the topical composition. Other suitable ranges are from 0.05 wt% to 2 wt% and from 0.1 wt% to 1 wt%.

[0051] In all embodiments of the invention, the use may be therapeutic or non-therapeutic. However, preferably, the use is non-therapeutic, including cosmetic uses.

[0052] The term "cosmetically acceptable carrier" (also referred to herein as a carrier) means all carriers / carriers conventionally used in topical cosmetic compositions, i.e., carriers / carriers suitable for topical application to the stratum corneum, possessing good aesthetic properties, compatibility with the active substances present in the composition, and not causing any unreasonable safety or toxicity issues. Such carriers are well known to those skilled in the art.

[0053] The exact amount of the carrier will depend on the actual level of the *Salix viminalis* extract, preferably *Salix flecherii* extract, and any other optional ingredients (e.g., other active ingredients) that a person skilled in the art would classify as different from the carrier.

[0054] In an advantageous embodiment, the local composition according to the invention comprises, based on the total weight of the local composition, about 50% to about 99%, preferably about 60% to about 98%, more preferably about 70% to about 98%, and particularly, about 80% to about 95% of a carrier.

[0055] In a particularly advantageous embodiment, the carrier also comprises at least 40% by weight, more preferably at least 50% by weight, and most preferably at least 55% by weight of water, for example, particularly about 55% by weight to about 90% by weight of water.

[0056] The local composition according to the invention is preferably prepared by blending / incorporating a Willowherb extract having all the definitions and preferences given herein, preferably a Willowherb freschaefera extract, with a cosmetically acceptable carrier.

[0057] The use of the *Salix viminalis* extract according to the present invention, preferably the *Salix frieshe* extract, can be therapeutic or non-therapeutic. Preferably, the use is non-therapeutic, i.e., for cosmetic applications. Therefore, preferably, in all embodiments, the topical composition is a cosmetic (non-therapeutic) composition.

[0058] As used in this application, the term "cosmetic composition" refers to, as in Cosmetic compositions as defined under the title “Kosmetika” in Lexikon Chemie, 10th edition, 1997, Georg Thieme Verlag Stuttgart, New York, and cosmetic compositions as disclosed in A. Domsch, "Cosmetic Compositions", Verlag für chemischeIndustrie (edited by H. Ziolkowsky), 4th edition, 1992.

[0059] Suitable compositions according to the invention are rinse-free or wash-off products, and include any products applied to the human body.

[0060] The composition may be in the form of a liquid, lotion, cream, foam, scrub, gel, soap bar, or toner, or applied by an apparatus or via a mask, cotton pad, or patch. Non-limiting examples of such compositions include leave-on lotions and creams, shampoos, conditioners, body gels, facial cleansers, shower gels, soap bars, antiperspirants, deodorants, hair removal products, lipsticks, foundations, mascaras, tanning creams, and sunscreens.

[0061] The preferred composition according to the invention is a no-rinse composition, more preferably in the form of an emulsion or gel.

[0062] The compositions of the present invention (including a carrier) may contain conventional adjuvants and additives, such as preservatives / antioxidants, fatty substances / oils, organic solvents, siloxanes, thickeners, softeners, emulsifiers, defoamers, soaps, detergents, aesthetic components (e.g., fragrances), surfactants, fillers, anionic polymers, cationic polymers, nonionic polymers or amphoteric polymers or mixtures thereof, propellants, acidifiers or alkalizers, dyes, colorants / dyes, abrasives, absorbents, chelating agents and / or multivalent chelating agents, essential oils, skin-feeling agents, astringents, pigments, or any other ingredients commonly formulated into such compositions.

[0063] According to the present invention, the compositions may further comprise other cosmetic active ingredients conventionally used in cosmetic compositions. Exemplary active ingredients include ultraviolet filters; agents for preventing or reducing inflammation; firming agents, moisturizers, soothing agents and / or energizing agents; and agents for improving elasticity and the skin barrier.

[0064] Examples of cosmetic excipients, diluents, adjuvants, additives, and active ingredients commonly used in the skin care industry that are suitable for use in the cosmetic compositions of the present invention are described, for example, in, but not limited to, the International Cosmetic Ingredient Dictionary & Handbook by Personal Care (http: / / www.personalcarecouncil.org / ), provided by the Personal Care Products Council, which is accessible online at INFO BASE (http: / / online.personalcarecouncil.org / jsp / Home.jsp).

[0065] The necessary amounts of the active ingredient, excipients, diluents, adjuvants, additives, etc., can be easily determined by a technician based on the desired product form and application. Additional ingredients may be added to the oil phase, aqueous phase, or added separately, as appropriate.

[0066] In some cases, the cosmetic active ingredients used in this invention may provide more than one benefit or act through more than one mode of action.

[0067] Of course, those skilled in the art will take care to select the optional additional ingredients, adjuvants, diluents and additives and / or their amounts such that the advantageous properties inherently associated with the combination according to the invention are not, or substantially not, adversely affected by one or more of the contemplated additions.

[0068] If the composition is an emulsion, particularly an oil-in-water (O / W), water-in-oil (W / O), silicone-in-water (Si / W), silicone-in-water (W / Si), oil-in-water-in-oil (O / W / O), water-in-oil-in-water (W / O / W) type or Pickering emulsion, the amount of oil phase present in such cosmetic emulsions is preferably at least 10% by weight based on the total weight of the composition, for example, in the range of 10% by weight to 60% by weight, preferably in the range of 15% by weight to 50% by weight, and most preferably in the range of 15% by weight to 40% by weight.

[0069] In one embodiment, the composition according to the invention is advantageously in the form of an O / W emulsion, said O / W emulsion comprising an oil phase dispersed in an aqueous phase in the presence of an O / W emulsifier. The preparation of such O / W emulsions is well known to those skilled in the art.

[0070] If the composition according to the invention is an O / W emulsion, it advantageously contains at least one O / W or Si / W type emulsifier selected from the following list: glyceryl stearate citrate, glyceryl stearate SE (self-emulsifying), stearic acid, stearate, polyglycerol-3-methylglucose distearate. Other suitable emulsifiers are phosphate esters and their salts, such as cetyl phosphate (e.g., from DSM Nutritional Products Ltd.). A) Diethanolamine cetyl phosphate (e.g., from DSM Nutritional Products Ltd.) DEA), cetyl phosphate (e.g., from DSM Naturitional Products Ltd.) K), sodium cetearyl sulfate, sodium glycerol oleate phosphate, hydrogenated vegetable glycerides phosphate, and mixtures thereof. Other suitable emulsifiers are polyalkylene glycol ethers, sorbitan oleate, sorbitan sesquioleate, sorbitan isostearate, sorbitan trioleate, cetearyl glucoside, dodecyl glucoside, decyl glucoside, sodium stearoyl glutamate, sucrose polystearate, and hydrated polyisobutylene. In addition, one or more synthetic polymers can be used as emulsifiers. For example, PVP eicosene copolymer, acrylate / C 10-30 Alkyl acrylate crosspolymers, and mixtures thereof.

[0071] The at least one O / W or Si / W type emulsifier is preferably used in an amount based on the total weight of the composition, from 0.5 wt% to 10 wt%, particularly in the range of 0.5 wt% to 6 wt%, for example, more particularly in the range of 0.5 wt% to 5 wt%, for example, most particularly in the range of 1 wt% to 4 wt%.

[0072] Particularly suitable O / W emulsifiers for use in the compositions according to the invention encompass phosphate ester emulsifiers, such as advantageously C8-10 alkyl ethyl phosphates, C9-15 alkyl phosphates, cetearyl alcohol polyether-2 phosphates, cetearyl alcohol polyether-5 phosphates, cetearyl alcohol polyether-8 phosphates, cetearyl alcohol polyether-10 phosphates, cetearyl phosphates, C6-10 alkanol polyether-4 phosphates, C12-15 alkanol polyether-2 phosphates, C12- 15-Alkyl alcohol polyether-3 phosphate, DEA-cetearyl alcohol polyether-2 phosphate, DEA-cetearyl phosphate, DEA-oleyl alcohol polyether-3 phosphate, cetyl phosphate potassium, decanol polyether-4 phosphate, decanol polyether-6 phosphate and lauryl alcohol polyether-4 phosphate and polyalkylene glycol ethers, such as, in particular, polyethylene stearyl ether, such as stearyl alcohol polyether-2 (Steareth-2) and stearyl alcohol polyether-21 (Steareth-21).

[0073] One particularly suitable class of O / W emulsifiers for use in the compositions according to the invention is polyalkylene glycol ethers. Particularly preferred O / W emulsifiers in all embodiments of the invention are stearyl ethers of polyethylene glycol, such most preferably stearyl alcohol polyether-2 (polyoxyethylene (2) stearyl ether) or stearyl alcohol polyether-21 (polyoxyethylene (21) stearyl ether) and mixtures thereof. Such polyalkylene glycol ether emulsifiers are, for example, commercially available at Croda under the trade name Brij.

[0074] The particularly preferred gels according to the invention comprise water, a thickener (e.g., preferably an acrylate thickener), and optional other ingredients. The water content in such gels is preferably >85% by weight, more preferably >90% by weight.

[0075] The topical compositions according to the invention advantageously contain a preservative. Particularly suitable preservatives in all embodiments of the invention are phenoxyethanol and ethylhexylglycerin, and mixtures thereof. When present, the preservative is used in an amount based on the total weight of the composition, preferably from 0.1% to 2% by weight, more preferably from 0.5% to 1.5% by weight.

[0076] The pH of the compositions according to the invention is generally in the range of 3 to 10, preferably in the range of 4 to 8, and most preferably in the range of 5 to 8. The pH can be readily adjusted as needed using suitable acids (e.g., citric acid) or bases (e.g., sodium hydroxide (e.g., aqueous solution), triethanolamine (TEA Care), tromethamine (Trizma base), and aminomethylpropanol (AMP-Ultra PC 2000) according to standard methods in the art.

[0077] The following examples are provided to further illustrate the compositions and effects of the present invention. These examples are merely illustrative and are not intended to limit the scope of the invention in any way. Example

[0078] In vivo studies were conducted to demonstrate the prebiotic effects of formulated *Salix frescoeflora* extract treatment on the facial microbiota.

[0079] Methods: In a 28-day in vivo placebo-controlled clinical study, a light gel formulation containing 3% *Salix frescoe* flower / leaf / stem extract, as listed in Table 1, was applied twice daily to the face. Sebum levels were measured before and after treatment to confirm the known sebum-reducing effect of *Salix frescoe* extract. Additionally, microbiota samples were collected from different facial sites (forehead, nasal alae, anterior cheek, lateral cheek, and chin) at the start of the study and 28 days after treatment. 16S rRNA sequencing was performed on the extracted DNA. Results from 16S rRNA sequencing were processed according to the “reference frames” method published by Morton et al. (Nat. Commun., 2019.10(1): p. 2719). This method allows for the assessment of microbiota changes and how these changes correlate with time and treatment. Differences were calculated using software called Songbird, starting with relative abundance data from 16S sequencing.

[0080] Table 1. Formulations

[0081]

[0082] Results: The differences in time and treatment are reported in Tables 2 and 3 below.

[0083] The “Sampling Location” column indicates the location where the facial microbiota sample was collected. The “Intercept” column indicates the starting point for each microorganism. A positive value indicates that the microorganism of interest was present on the skin to some extent prior to evaluation. The “Treatment” column indicates whether and to what extent a particular microorganism is associated with the active formulation treatment. The reported values ​​are the result of a difference made by the software using a placebo sample as a reference. Therefore, a positive value indicates a positive correlation with the active formulation treatment, while a negative value indicates a negative correlation with the treatment. The “Time” column indicates how a particular microorganism is associated with time, but not with treatment. A positive value indicates that the microorganism tends to increase its abundance over time.

[0084] First, we identified several species that showed a positive correlation with treatment with *Gynostemma flecheri* extract compared to placebo treatment (i.e., control) and were also known to have positive effects on the human skin microbiome. This is reflected in the positive difference values ​​in the "Treatment" column of Table 2, therefore we consider the treatment to be beneficial to the growth of the selected species.

[0085] Table 2: "Reference Frame" Analysis of Positively Regulatory Species

[0086] Sampling location intercept treat time Species cheek 1.27751447873 0.228874916708 -0.544504983788 Staphylococcus aureus Side cheek 1.00349654700 0.4281320781321 -0.1412418596446 Staphylococcus aureus forehead 2.77272038213 0.561305068256 0.1242964515655 Staphylococcus epidermidis jaw 2.57411628058 0.535163912442 0.1115589713056 Staphylococcus epidermidis cheek 1.841159364756 1.114582757032 -1.101744516259 Crowded Corynebacterium nose 1.821209192276 0.882371737239 -0.2124683622470 Propionibacterium granulosum jaw 0.886907866506 0.645104977753 -0.2319768319527 Propionibacterium granulosum forehead 1.760498330510 1.141440681936 -0.533702102843 Yunnan Micrococcus nose 0.672707796096 0.596773041961 -0.0134971428862 Yunnan Micrococcus Side cheek 1.871380229254 0.3884648294062 -0.365194153040 Yunnan Micrococcus jaw 0.767720511465 0.622249934342 -0.3413930411140 Yunnan Micrococcus

[0087] Furthermore, we identified species that were negatively correlated with treatment with *Salix frescoe* extract (compared to placebo) and were also known to have negative or potentially pathogenic effects on human skin. This is reflected in the negative difference values ​​in the "Treatment" column of Table 3, therefore we conclude that the treatment inhibited or reduced the growth of the selected species.

[0088] Table 3: "Reference Frame" Analysis of Negatively Regulatory Species

[0089]

[0090]

[0091] Therefore, in summary, considering the treatment-based outcomes for species listed in group-outcome format in Tables 2 and 3, it is clear that for the skin, the microbiota shifted towards a healthier composition, as the growth and relative abundance of species positively correlated with skin health were promoted, and the treatment reduced species with negative effects on the skin or pathogenic potential. Thus, we can conclude that the treatment promoted both specific regulation of species growth and inhibition, leading to a positive rebalancing of the facial microbiota, which can be attributed to the prebiotic effect of the application of *Salix frescoe* extract.

Claims

1. The non-therapeutic use of *Salix flecheri* extract as a prebiotic when applied to the external surface of the human body, wherein the use as a prebiotic is characterized by increasing the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium pumilus in the skin microbiota of the individual in need, compared with a control.

2. The use according to claim 1, wherein the *Willow Leaf Frischii* extract is an aqueous extract obtained from the flowers / leaves and stems of the corresponding *Willow Leaf Frischii* plant.

3. The use according to claim 1, wherein the outer surface of the human body is skin.

4. The use according to claim 1, wherein the outer surface of the human body is at least one of the scalp, including hair, and the armpit.

5. The use according to claim 1, wherein the Willowherb flechery extract is applied in the form of a topical composition comprising an effective amount of the Willowherb flechery extract and a cosmetically acceptable carrier.

6. The use according to claim 5, wherein the topical composition comprises 0.001% to 10% by weight of the *Willow Leafy Greens* extract based on the total weight of the topical composition.

7. The use according to claim 5 or 6, wherein the topical composition is a no-rinse composition.

8. The use according to claim 7, wherein the rinse-free composition is in the form of an emulsion or gel.

9. The use according to claim 1, wherein the use as a prebiotic is characterized by reducing, compared with a control, the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus capitella, Corynebacterium tuberculosis, Lawsonia clevelandii, Staphylococcus aureus, and Corynebacterium cropensteine ​​in the skin microbiota of the individual in need.

10. A method for increasing the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium pulvinatum in an individual’s skin microbiota, the method comprising the step of topically applying an effective amount of Willowleaf frescoe extract to the desired external surface of the body.

11. The method of claim 10, wherein the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus capitulata, Corynebacterium tuberculosis, Lawsonia clevelandii, Staphylococcus aureus, and Corynebacterium cropensteinii is simultaneously reduced.

12. A method for providing protection to the outer surface of a human body by increasing the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium claustrophosum, the method comprising the step of applying a Willowleaf frechetum extract to the outer surface.

13. The use of *Salix flecheri* extract for increasing the differential abundance of at least one microorganism selected from the group consisting of Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus yunnanensis, Propionibacterium granulosum, and Corynebacterium pumilus in the skin microbiota of individuals in need.

14. The use according to claim 13, wherein the differential abundance of Staphylococcus capitella, Corynebacterium tuberculosis, Lawsonia clevelandii, Staphylococcus aureus, and Corynebacterium cropensteinii is reduced.

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